Biological Sequence Retrieval

Retrieve DNA, RNA, and protein sequences with proper disambiguation and cross-database handling.

IMPORTANT: Always use English terms in tool calls. Only try original-language terms as fallback. Respond in the user's language.

LOOK UP DON'T GUESS: Never assume accession numbers or sequence versions. Always retrieve and verify from NCBI or ENA.

Domain Reasoning

Sequence quality hierarchy: RefSeq (NM_/NP_ = curated) > RefSeq predicted (XM_/XP_) > GenBank (submitted). Prefer the MANE Select transcript for human canonical isoforms. Check version numbers -- annotations improve across versions.

Workflow

Phase 0: Clarify (if needed) → Phase 1: Disambiguate Gene/Organism → Phase 2: Search & Retrieve → Phase 3: Report

Phase 0: Clarification (When Needed)

Ask ONLY if: gene exists in multiple organisms, sequence type unclear, or strain matters. Skip for: specific accessions, clear organism+gene combos, complete genome requests with organism.

Phase 1: Gene/Organism Disambiguation

Accession Type Decision Tree

| Prefix | Type | Use With | |--------|------|----------| | NC_/NM_/NR_/NP_/XM_ | RefSeq | NCBI only | | U/M/K/X/CP/NZ_ | GenBank | NCBI or ENA | | EMBL format | EMBL | ENA preferred |

CRITICAL: Never try ENA tools with RefSeq accessions -- they return 404.

Identity Checklist

• Organism confirmed (scientific name)
• Gene symbol/name identified
• Sequence type determined (genomic/mRNA/protein)
• Accession prefix identified for tool selection

Phase 2: Data Retrieval (Internal)

Retrieve silently. Do NOT narrate the search process.

python
# Search NCBI Nucleotide
result = tu.tools.NCBI_search_nucleotide(
    operation="search", organism=organism, gene=gene,
    strain=strain, keywords=keywords, seq_type=seq_type, limit=10
)

# Get accessions from UIDs
accessions = tu.tools.NCBI_fetch_accessions(operation="fetch_accession", uids=result["data"]["uids"])

# Retrieve sequence (FASTA or GenBank format)
sequence = tu.tools.NCBI_get_sequence(operation="fetch_sequence", accession=accession, format="fasta")

# ENA alternative (non-RefSeq accessions only)
entry = tu.tools.ena_get_entry(accession=accession)
fasta = tu.tools.ena_get_sequence_fasta(accession=accession)

Fallback Chains

| Primary | Fallback | Notes | |---------|----------|-------| | NCBI_get_sequence | ENA (if GenBank format) | NCBI unavailable | | ena_get_entry | NCBI_get_sequence | ENA doesn't have RefSeq | | NCBI_search_nucleotide | Try broader keywords | No results |

Phase 3: Report Sequence Profile

Present as a Sequence Profile Report. Hide search process. Include:

1. Search Summary: query, database, result count
2. Primary Sequence: accession, type (RefSeq/GenBank), organism, strain, length, molecule, topology, curation level
3. Sequence Preview: first lines of FASTA (truncated)
4. Annotations Summary: CDS/tRNA/rRNA/regulatory feature counts (from GenBank format)
5. Alternative Sequences: ranked by relevance and curation, with ENA compatibility
6. Cross-Database References: RefSeq, GenBank, ENA/EMBL, BioProject, BioSample
7. Download Options: FASTA (for BLAST/alignment), GenBank (for annotation)

Curation Level Tiers

| Tier | Prefix | Description | |------|--------|-------------| | RefSeq Reference (best) | NC_, NM_, NP_ | NCBI-curated, gold standard | | RefSeq Predicted | XM_, XP_, XR_ | Computationally predicted | | GenBank Validated | Various | Submitted, some curation | | GenBank Direct | Various | Direct submission | | Third Party | TPA_ | Third-party annotation |

Reasoning Framework

Sequence quality: Prefer RefSeq over GenBank. Check version numbers. Sequences with "PREDICTED" in definition are not experimentally validated.

Accession guidance: RefSeq = NCBI-only. GenBank = mirrored in ENA/EMBL. Default to RefSeq mRNA (NM_) for human/model organisms; most complete genome assembly for microbial queries.

Cross-database reconciliation: Same sequence may have different accessions (e.g., GenBank U00096 = RefSeq NC_000913 for E. coli K-12). Always report both when available. Discrepancies between GenBank/RefSeq typically indicate RefSeq curation corrected submission errors.

Synthesis Questions

1. What is the highest-quality accession available?
2. Are there alternative accessions in other databases?
3. What is the annotation completeness?
4. Is the sequence from the expected organism/strain?
5. What download format suits the user's downstream analysis?

Error Handling

| Error | Response | |-------|----------| | "No search criteria provided" | Add organism, gene, or keywords | | "ENA 404 error" | Likely RefSeq -- use NCBI only | | "No results found" | Broaden search, check spelling, try synonyms | | "Sequence too large" | Note size, provide download link instead |

Tool Reference

NCBI Tools: NCBI_search_nucleotide (search), NCBI_fetch_accessions (UID→accession), NCBI_get_sequence (retrieve) ENA Tools (GenBank/EMBL only): ena_get_entry (metadata), ena_get_sequence_fasta (FASTA), ena_get_entry_summary (summary)

Search Parameters Reference

NCBI_search_nucleotide: operation="search", organism (scientific name), gene (symbol), strain, keywords, seq_type (complete_genome/mrna/refseq), limit

NCBI_get_sequence: operation="fetch_sequence", accession, format (fasta/genbank)